Patch-clamp studies in human macrophages: Single-channel and whole-cell characterization of two K+ conductances

1988 ◽  
Vol 103 (1) ◽  
pp. 55-66 ◽  
Author(s):  
Elaine K. Gallin ◽  
Leslie C. McKinney
Keyword(s):  
Patch Clamp ◽  
Single Channel ◽  
Whole Cell ◽  
Human Macrophages ◽  
Cell Characterization

scholarly journals The Tyrosine Kinase p56lck Mediates Activation of Swelling-induced Chloride Channels in Lymphocytes

1998 ◽  
Vol 141 (1) ◽  
pp. 281-286 ◽  
Author(s):  
Albrecht Lepple-Wienhues ◽  
Ildikò Szabò ◽  
Tilmann Laun ◽  
Nubia Kristen Kaba ◽  
Erich Gulbins ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Tyrosine Kinase ◽  
Single Channel ◽  
Regulatory Volume Decrease ◽  
Cell Swelling ◽  
Osmotic Swelling ◽  
Whole Cell ◽  
Herbimycin A ◽  
Unitary Conductance ◽  
Human T Cell

Osmotic cell swelling activates Cl− channels to achieve anion efflux. In this study, we find that both the tyrosine kinase inhibitor herbimycin A and genetic knockout of p56lck, a src-like tyrosine kinase, block regulatory volume decrease (RVD) in a human T cell line. Activation of a swelling-activated chloride current (ICl−swell) by osmotic swelling in whole-cell patch-clamp experiments is blocked by herbimycin A and lavendustin. Osmotic activation of ICl−swell is defective in p56lck-deficient cells. Retransfection of p56lck restores osmotic current activation. Furthermore, tyrosine kinase activity is sufficient for activation of ICl−swell. Addition of purified p56lck to excised patches activates an outwardly rectifying chloride channel with 31 pS unitary conductance. Purified p56lck washed into the cytoplasm activates ICl−swell in native and p56lck-deficient cells even when hypotonic intracellular solutions lead to cell shrinkage. When whole-cell currents are activated either by swelling or by p56lck, slow single-channel gating events can be observed revealing a unitary conductance of 25–28 pS. In accordance with our patch-clamp data, osmotic swelling increases activity of immunoprecipitated p56lck. We conclude that osmotic swelling activates ICl−swell in lymphocytes via the tyrosine kinase p56lck.

Patch-clamp study of single-channel and whole-cell K+ currents in guinea pig pancreatic acinar cells

1988 ◽  
Vol 255 (3) ◽  
pp. G275-G285 ◽  
Author(s):  
K. Suzuki ◽  
O. H. Petersen
Keyword(s):  
Patch Clamp ◽  
Guinea Pig ◽  
Single Channel ◽  
Pancreatic Acinar Cells ◽  
Intact Cells ◽  
Membrane Area ◽  
Pancreatic Acini ◽  
Whole Cell ◽  
Single Channel Currents ◽  
K Currents

K+ channels in the plasma membrane of isolated guinea pig pancreatic acini were studied by patch-clamp single-channel and whole-cell current recording techniques. Three types of K+-permeable pores were found in excised patch experiments: Ca2+-activated nonselective cation channels with a unit conductance of approximately 25 pS that could be inhibited by ATP acting on the membrane inside, and two kinds of Ca2+- and voltage-activated K+-selective channels with unit conductances (in symmetrical K+-rich solutions) of about 200 and 30 pS, respectively. In intact cells, pentagastrin activation of currents through the 30 pS K+-selective pores was demonstrated. In these experiments pentagastrin was added to the bath solution and had no direct contact with the electrically isolated membrane area from which the single-channel currents were recorded, suggesting that the activation is mediated via an intracellular messenger system. Pentagastrin stimulation of voltage-gated K+ currents was also observed in whole-cell recording experiments. Results from these experiments suggest that in the stimulated condition the membrane electrical properties were dominated by the 30 pS K+-selective channels.

Activation of Cl− currents by intracellular chloride in fibroblasts stably expressing the human cystic fibrosis transmembrane conductance regulator

1993 ◽  
Vol 71 (9) ◽  
pp. 645-649 ◽  
Author(s):  
Xiaodong Wang ◽  
Yoshinori Marunaka ◽  
Ludwik Fedorko ◽  
Sascha Dho ◽  
J. Kevin Foskett ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Cyclic Amp ◽  
Patch Clamp ◽  
Single Channel ◽  
Fibroblast Cell ◽  
Mouse Fibroblast ◽  
Transmembrane Conductance Regulator ◽  
Transmembrane Conductance ◽  
Whole Cell ◽  
Cystic Fibrosis Transmembrane

The Cl− conductance of a mouse fibroblast cell line (LTK− cells) that was stably transfected with the human CFTR (cystic fibrosis transmembrane conductance regulator) complementary DNA was studied. Single Cl− channel activity was observed only after treatment of the cells with forskolin, the single-channel conductance being 6.2 ± 0.2 pS with a linear current–voltage relationship. In CFTR+ cells, the whole-cell current at +90 mV increased from 7.3 ± 2.7 pA/pF (n = 12) to 46.1 ± 11.2 pA/pF (n = 5) after addition of dibutyryl-cyclic AMP (10−4 M) to the bath. Increasing the intracellular Cl− concentration to 150 mM activated linear Cl− currents in the absence of cyclic AMP in CFTR+ (n = 42) but not in CFTR− cells (n = 4). Similar Cl− current was also activated by high intracellular I− concentration. These results indicate that the CFTR-induced Cl− conductance in LTK− cells can be activated by either cyclic AMP or high intracellular halide concentrations.Key words: cystic fibrosis transmembrane conductance regulator (CFTR), chloride channel, cyclic AMP, whole-cell patch clamp, single-channel patch clamp.

scholarly journals Characterization of ryanodine receptor type 1 single channel activity using “on-nucleus” patch clamp

Cell Calcium ◽  
2014 ◽  
Vol 56 (2) ◽  
pp. 96-107 ◽  
Author(s):  
Larry E. Wagner ◽  
Linda A. Groom ◽  
Robert T. Dirksen ◽  
David I. Yule
Keyword(s):  
Patch Clamp ◽  
Ryanodine Receptor ◽  
Single Channel ◽  
Receptor Type ◽  
Channel Activity ◽  
Single Channel Activity

scholarly journals Properties of single-channel and whole cell Cl− currents in guinea pig detrusor smooth muscle cells

2019 ◽  
Vol 316 (5) ◽  
pp. C698-C710 ◽  
Author(s):  
Viktor Yarotskyy ◽  
John Malysz ◽  
Georgi V. Petkov
Keyword(s):  
Smooth Muscle ◽  
Patch Clamp ◽  
Guinea Pig ◽  
Smooth Muscle Cells ◽  
Single Channel ◽  
Muscle Cells ◽  
Niflumic Acid ◽  
Detrusor Smooth Muscle ◽  
Whole Cell ◽  
Whole Cell Patch Clamp

Multiple types of Cl− channels regulate smooth muscle excitability and contractility in vascular, gastrointestinal, and airway smooth muscle cells. However, little is known about Cl− channels in detrusor smooth muscle (DSM) cells. Here, we used inside-out single channel and whole cell patch-clamp recordings for detailed biophysical and pharmacological characterizations of Cl− channels in freshly isolated guinea pig DSM cells. The recorded single Cl− channels displayed unique gating with multiple subconductive states, a fully opened single-channel conductance of 164 pS, and a reversal potential of −41.5 mV, which is close to the ECl of −65 mV, confirming preferential permeability to Cl−. The Cl− channel demonstrated strong voltage dependence of activation (half-maximum of mean open probability, V0.5, ~−20 mV) and robust prolonged openings at depolarizing voltages. The channel displayed similar gating when exposed intracellularly to solutions containing Ca2+-free or 1 mM Ca2+. In whole cell patch-clamp recordings, macroscopic current demonstrated outward rectification, inhibitions by 4,4′-diisothiocyano-2,2′-stilbenedisulfonic acid (DIDS) and niflumic acid, and insensitivity to chlorotoxin. The outward current was reversibly reduced by 94% replacement of extracellular Cl− with I−, Br−, or methanesulfonate (MsO−), resulting in anionic permeability sequence: Cl−>Br−>I−>MsO−. While intracellular Ca2+ levels (0, 300 nM, and 1 mM) did not affect the amplitude of Cl− current and outward rectification, high Ca2+ slowed voltage-step current activation at depolarizing voltages. In conclusion, our data reveal for the first time the presence of a Ca2+-independent DIDS and niflumic acid-sensitive, voltage-dependent Cl− channel in the plasma membrane of DSM cells. This channel may be a key regulator of DSM excitability.

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